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KMID : 0380020200350030199
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Korean Journal of Biotechnology and Bioengineering
2020 Volume.35 No. 3 p.199 ~ p.207
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Comparison of Physicochemical Factors Influencing the Production of Fibrinolytic Enzymes from Bacillus amyloliquefaciens G-13 in Fibrin Zymographic Gel
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Lee Hyun-Ho
Cha Min-Ji
Oh Kye-Heon
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Abstract
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Our previous research demonstrated the characteristics and fibrinolytic activities of NaCl- and capsaicin-resistant bacterium Bacillus amyloliquefaciens G-13, isolated from mustard leaf kimchi [1]. In this study, we extended the work to the comparison of various physicochemical factors (e.g., temperature, pH, NaCl, capsaicin, metal ions, and inhibitors) influencing the production of fibrinolytic enzyme by strain G-13. Initially, a time course of the fibrinolytic activity of strain G-13 was measured by the fibrin plate assay, and after 84 hours of incubation, the maximum fibrinolytic activity was about 3.42 times that of plasmin used as the standard. Through a fibrin zymographic assay, four bands (23, 34, 45, and 67 kDa) of fibrinolytic enzyme from strain G-13 were compared in fibrin zymographic gels. The strain could grow and produce the fibrinolytic enzyme in the presence of NaCl (1-10%) or capsaicin (0-300 μg/mL), respectively. The optimum pH and temperature for the enzymatic activity were 8 and 35oC. In the study of the effects of metal ions on fibrinolytic activity, K+, Ca2+, and Mg2+ ions increased the activity and maintained it overall, but Fe3+, Zn2+, Ba2+, and Hg2+ ions rapidly decreased the activity of the fibrinolytic enzyme. The enzyme was completely inhibited by phenylmethanesulfonyl fluroide (PMSF), suggesting that it was a serine protease. These results provide important clues for understanding the characteristics of fibrinolytic bacteria by confirming the effects of B. amyloliquefaciens G-13 on various physicochemical factors influencing the production of fibrinolytic enzymes through fibrin zymography.
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KEYWORD
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Bacillus amyloliqeufaciens G-13, fibrinolytic enzyme, fibrin zymograpy
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